WO2012045853A1

WO2012045853A1 - Illumination chamber for raman spectroscopy

Info

Publication number: WO2012045853A1
Application number: PCT/EP2011/067533
Authority: WO
Inventors: Philippe Bourget; Bruno Cassard
Original assignee: Assistance Publique - Hopitaux De Paris
Priority date: 2010-10-07
Filing date: 2011-10-07
Publication date: 2012-04-12
Also published as: FR2965920B1; PT2625506E; EP2625506A1; PL2625506T3; EP2625506B1; ES2511165T3; FR2965920A1; US20130301043A1; CA2813649A1; JP2013539045A; DK2625506T3

Abstract

The invention relates to a chamber for analyzing a sample by means of Raman spectroscopy, characterized in that the chamber is provided in the form of an opaque parallelepiped having: (a) on one of the vertical walls thereof, a hatch for introducing a sample; (b) on the bottom wall thereof, a first opening, and means (4) for attaching a light source, for providing excitation radiation, and a lens (22) enabling the convergence of the excitation radiation so as to define a focal point for the excitation radiation; and (c) a second opening and means for attaching a sensor enabling the Raman scattered light to be detected on a vertical wall or the bottom wall, said attachment means (4) including adjustment means (8) for varying said focal point of the excitation radiation along a vertical axis, so as to enable an adjustment of the height of said focal point within the analysis chamber.

Description

ILLUMINATION CHAMBER FOR RAMAN SPECTROSCOPY

The invention relates to the field of controlling the nature of samples by Raman spectroscopy. In particular, the invention is particularly useful for the analytical quality control (CQA) of products prepared extemporaneously in hospital pharmacies.

The "certification of Therapeutic Objects (OT)" via the concept of in situ release CQA, aims in fact to guarantee the parameters, identity, purity and concentration of the molecules of interest. These products may be of high added value (for example monoclonal antibodies), or may require precautions for use (for example cytotoxicity for products intended for chemotherapy).

In the best case, this analysis must be rapid (result in minutes and before infusion to the patient), without any consequences for the sample, and without it being necessary to take the sample for the purpose of control (certain samples can not be removed without being destroyed).

In this respect, Raman spectroscopy has a number of advantages:

- respect for the integrity of the therapeutic object: no withdrawal or destruction of sample

- direct / extemporaneous analysis and through the walls of the therapeutic object

- analyzes conducted under ambient conditions

simultaneous obtaining of the spectral signatures of the envelopes, vehicles, molecules of interest

- high speed (<2 min) / functional flexibility compatible with the principles of release CQA i.e. just before administering the OT to the patient. This rapid response also makes it possible to re-prepare an OT declared "non-compliant"

- no use of consumables or generation of production waste: environmental preservation and sustainable development

- remarkable specificity (requires analytical expertise)

- relatively simple implementation, compared to the reference separation methods - relatively low financial investment (tens of thousands of euros)

- low maintenance cost: few moving parts, motors ...

- size of the equipment making it usable in CPCI (Central Hospital Production or not Injectables or Chemotherapies)

- potentially large scope

- Water practically does not diffuse in Raman spectroscopy, making it useful as a "control spectrum"

The use of Raman technology also makes it possible to identify, and still without intrusion, the vehicle (NaCl 0.9%, G5%, water ppi ...) carrying the molecule of interest.

The principle of Raman spectroscopy is as follows: the radiations of a powerful monochromatic (laser) source are conducted in an optical fiber, and focused on the sample to be analyzed, thus provoking its excitation. The light scattered by the sample is collected by a sensor and then fed through an optical fiber to the separator (monochromator). Coupled with a detector it then provides data on the sample that need only be processed by computer. In general, the scattered light is collected either at 180 ° or at 90 °.

By comparison with reference spectra, it is thus possible to characterize the nature of the product present in the sample, its concentration, as well as the presence of any impurities.

This is a local measurement technique: the laser beam is focused on a small part of the sample, and allows to study the properties of this sample on a volume of a few cubic microns.

This technology has already been used for the analysis of capsules, or injectable solutions, and especially directly in plastic bottles.

The particular problem that arises in a hospital environment for the implementation of the CQA is the diversity of therapeutic objects to study, both in their composition and in the nature and shape of the container. Indeed, these therapeutic objects, when it comes to injectable solutions can be in syringes (of various volumes), infusion bags, bottles, beakers or similar containers, ampoules, or infusers portable.

Portable infusers are lightweight, disposable devices to enable slow, continuous infusion of chemotherapy. The product of Chemotherapy is contained in a flexible balloon reservoir, which is itself placed inside a rigid envelope.

The wide variety of therapeutic objects and their geometry requires a system to apply the Raman spectroscopy method to these objects, easily. In addition, hospital pharmacies preparing multiple Therapeutic Objects, it is necessary to be able to substitute one object to another quickly, without the setting of the excitation light source is difficult to perform. Finally, operators should also be protected from any exposure to the laser beam or scattered light.

The invention thus relates to a chamber for the analysis of a sample by Raman spectroscopy, characterized in that it is in the form of an opaque parallelepiped, presenting

(a) a trap for introducing a sample, on one of its vertical walls,

(b) a first port and means (4) for attaching a light source to provide excitation radiation and a lens for convergence of the excitation radiation to define a focal point of the radiation of excitation, on the lower wall

(c) a second port and sensor attachment means for detecting the Raman scattered light, on a vertical wall or on the bottom wall

said fixing means (4) comprising adjustment means (8) for varying said focus point of the excitation radiation on a vertical axis to allow adjustment of the height of said focusing point within the chamber analysis. The terms "vertical" and "lower" are related to how the chamber is preferentially set up when it is used to analyze a sample. In fact, this clearly means that the sample introduction flap is located on a first wall. The first port and the means (4) for attaching a light source to provide excitation radiation and a lens (22) for convergence of the excitation radiation to define a focus point of the excitation radiation are located on a second wall which is perpendicular to the first wall.

The second orifice and the means for fixing a sensor for detecting Raman scattered light are located on a wall, which is not parallel (vis-à-vis) to the second wall.

In this embodiment, the light source (laser head) and the lens are located on the lower wall of the chamber according to the invention. The light is sent from the bottom to the top. This embodiment is preferred because it makes it possible to characterize the sample which is placed on the floor of the lower wall, above the first orifice.

However, in other embodiments which are also part of the invention, it is possible to implant the laser head on a sidewall (the first orifice then being in a side wall of the chamber), this actually being to "turn" the room during use. Several laser heads can also be implanted for the acquisition of Raman data in different points of the sample. In this case, however, it should be checked that there is no interference between the different detectors, so that each detects the scattered light after excitation by a specific head.

As we have seen, Raman spectroscopy amounts to collecting and analyzing the light scattered by the sample after excitation, generally at 90 ° or 180 ° from the axis of the excitation laser beam.

In a preferred embodiment, the second port is coincident with the first port. In this embodiment, the scattered light is collected at 180 °. In this embodiment, the fixing means (4) and the fixing means of said sensor are also merged. They then contain two optical fibers (one bringing the light emitted via the laser head, and the second conveying the light scattered by the sample towards the monochromator and the detector).

Due to the great variability of the Therapeutic Objects to be tested, and in particular the thickness of the envelopes, it is necessary to be able to move the focusing point of the excitation light, so that it is located in the sample, at approximately 1 cm above the envelope of the Therapeutic Object. This distance makes it possible in particular to avoid the "edge effects" related to the envelope. As mentioned above, the laser (parallel monochromatic light source) is coupled to a lens allowing the convergence of the light beams in a small volume of the sample that is to be analyzed. The distance between the lens and the point of convergence is known to the operator, and is expressed by the focal length of the lens.

In one embodiment, the fastening means (4) enable the light source (the laser head) and the lens to be moved together.

In another embodiment, the laser head is fixed relative to the bottom wall of the analysis chamber, and only the lens is movable on the vertical axis.

In a preferred embodiment, said fixing means (4) of said light source and the lens comprise a light-tight sleeve attached to the bottom wall of the chamber.

In a particular embodiment, this sleeve is screwed to the bottom wall of the illumination chamber. In this embodiment, it is possible to machine recesses in the sole, to introduce the screws, in order to maintain the flatness of the sole after fixing the sleeve.

The light tightness of the sleeve is achieved by making it in an opaque material (such as polycarbonate). This allows both to avoid the "contamination" of the light beam by the ambient light and to protect the operators.

In a particular embodiment, the adjustment means of the moving assembly (which is the light source and the lens when these two elements are integral or the lens alone) comprise a millimeter screw. The use of a millimetric screw and can move on the elements to be movable on the vertical axis very accurately. It is thus possible to use a millimetric screw allowing positioning to the hundredth of a millimeter. The principle of the millimeter screw is to induce a translational movement from a rotational movement of the screw. The thread allows to define the precision of the screw (displacement in translation for each turn of screws).

In a preferred embodiment, this screw is connected to a counter for knowing the displacement of the screw, equal to that of the moving assembly, and therefore the height of the focusing point within the chamber according to the invention. By way of illustration, it is assumed that the focal length of the lens is 20 mm, and that the moving assembly is flush with the floor of the chamber at point 0 of the counter. This implies that the focal point of the light is 20 mm above the floor.

If one seeks to analyze a sample present in a plastic envelope 1 mm thick, we will seek to achieve the focus to 1 1 mm above the sole (about 1 cm in the sample). We must therefore lower the moving object by 9 mm.

If the sample is present in a syringe with a thickness of

3 mm, the moving object is lowered by 7 mm.

Those skilled in the art thus determine the adjustment to be made to the moving object according to the focal length of the lens, and the nature of the therapeutic object, and in particular the thickness of the envelope.

The chamber according to the invention (Raman illumination chamber) can be fixed on a frame comprising feet.

This frame may also include means for adjusting the height of the chamber relative to the frame (for example adjustable feet on a screw jack). In this case, it may be advantageous for the chamber to carry means for measuring the flatness, such as a spirit level, to ensure that the sole is perfectly horizontal.

The walls of the chamber are machined in any opaque material. It should be noted that the different walls can be in different materials. In a preferred embodiment, is used, at least for the sole, a material to ensure the best possible flatness, avoiding the maximum relief irregularities on the sole. It is thus possible to use polycarbonate or carbon material. The thickness of the walls of the chamber according to the invention is defined by those skilled in the art, is generally greater than 0.5 cm and generally does not exceed a few centimeters. It is thus possible to use carbon walls with a thickness of 1 cm.

The opacity of the chamber makes it possible to avoid measurement interference that could be due to ambient light during the implementation of the Raman spectroscopy measurement. It also helps protect operators. The sample introduction flap in the chamber according to the invention can be of any kind. However, it is important that it preserves the opacity of the room once closed. We can therefore provide a seal to do this.

The hatch can slide in a guillotine along a side wall (top to bottom or bottom to top). It can also be opened via a system of hinges, or a rack (motorized or not). Reminder systems (such as springs) can also be envisaged to assist the operator in closing the hatch after placing the sample. It can be fixed in closed position by any means (magnet, hook ...).

The invention also relates to a method of analyzing a sample present in an envelope comprising the steps of

(a) placing said sample on the floor of the chamber according to the invention, vertically above said first orifice and facing said second orifice,

(b) actuating the adjusting means (8) to position the focusing point of the excitation radiation in the sample about 1 cm above said envelope,

(c) emitting said excitation radiation and detecting Raman scattered light. It is clear that the excitation radiation is emitted via a light source fixed on the fixing means (4).

The detection of the Raman scattered light is obviously carried out by means of a sensor fixed on the means for fixing a sensor of said chamber.

As has been seen, this method is particularly useful when said sample is a liquid solution, and said envelope is chosen from a syringe, an infusion bag, a bottle, a beaker or similar container, a bulb, an infuser portable.

In another embodiment, the method is implemented with a sample that is solid, said envelope being chosen from a capsule, a capsule and a solid coating.

However, it is also possible to implement this method on solid samples without an envelope, and in particular tablets. This embodiment is also included within the scope of the invention. In one embodiment, the excitation radiation is emitted for a duration of between 1 second and one minute.

The duration of excitation is defined by the operator and depends in particular on the power of the laser, and the nature of the sample to be analyzed.

In order to increase the quality of the analysis, the sample is often subjected to several excitation radiations, and the data collected for each excitation is averaged.

It should be noted that the present invention also covers the use of Raman spectroscopy, in general, to characterize the contents of a portable brewer.

The invention also covers the use of Raman spectroscopy, in general, for characterizing an anticancer, as described in the examples. Description of Drawings

Figure 1 shows a sectional view of an embodiment of fixing and adjusting means according to the invention. The fixing means of the light source (4), in the form of a sleeve, and the adjustment means (8) in the form of a millimetric screw are visible. A hole (14) for fixing with a screw is also visible.

FIG. 2 represents a view from above of the means illustrated in FIG. 1. The three screw holes (14) and the location of the lens (22) are clearly visible. The light source is located behind the lens (22). The sensor may also be located at the same location as the lens (22).

Figure 3 shows a general view of fixing means (4) and adjustment (8) applicable according to the invention. We see the millimetric screw (8) (knob and screw thread) and a sleeve (4) containing the laser head, the lens (22) and optionally the sensor. The sheath of the optical fiber (15) allowing the arrival of light into the laser head from the power source is also shown. It may also contain a second optical fiber for transferring the scattered light received by the sensor to the monochromator and detector.

Figure 4 shows a chamber according to the invention. The fixing means (4) and setting (8), fixed on the lower wall of the chamber are shown, as well as the optical fiber sheath (15). Examples

Example I -Application to oxazaphosphorines (ifosfamide)

Operating mode

Material: Kaiser Optical® 785 nm RXN1 Raman Spectrometer

Laser: 785 nm - power 450 mW

Acquisition time: 1 min (room)

• 5 range points (0 - 1 - 5 - 10 - 15 - 20 mg / mL) (3x)

• 3 CQs (2 - 8 - 16 mg / mL) (6x)

· 3 infusion bags (2 - 4 - 6 mg / mL) (3x)

• Two dilution solvents: NaCl 0.9% and G 5%

Validation on 3 days

HPLC protocol: identical (same samples) Results

A linear peak intensity range is obtained as a function of concentration (1 to 20 mg / mL); r ² > 0.99 (n = 9); significant slope (P <0.001 - F test)

The variation of the slopes of the standard ranges <5%, and one observes a reproducibility of the data intra-day and inter-day.

These results show that this technology is applicable to oxazaphosphorines.

Example II - Application to anthracyclines (ifosfamide)

The same material as in Example 1 is used.

Application of Raman Spectroscopy (SR) to CQA of doxorubicin and epirubicin syringes, vehicle NaCl 0.9% or Glucose 5%.

These molecules exhibit an epimerization: inversion of steric configuration of the 4th carbon of the carbohydrate fraction and UV spectra: identical (Amax 254 nm)

The results obtained show that Raman spectroscopy makes it possible to measure the concentration of the two products in solution, whatever the vehicle used, with a good linearity.

In addition, two characteristic wavelengths (465 cm- ¹ and 350 cm- ¹ ) are observable for doxorubicin, making it possible to differentiate it from epirubicin. The nature of the spectra observed also makes it possible to differentiate the vehicle used.

Example III - Application to portable brewers

The same material as in Example 1 is used.

The problem of portable brewers is that it is impossible to take the sample once introduced into the infuser without destroying the brewer. Moreover, the presence of two envelopes surrounding the sample decreases the strength of the excitation signal (each envelope absorbing part of the excitation energy). It has been shown that despite the presence of both envelopes, Raman spectrometry allows to qualify the nature of the product contained in portable brewers.

Claims

Chamber for analysis of a sample by Raman spectroscopy, characterized in that it is in the form of an opaque parallelepiped, presenting

(a) on one of its vertical walls, a trap for introducing a sample,

(b) on the bottom wall, a first port and means (4) for attaching a light source to provide excitation radiation and a lens (22) for convergence of the excitation radiation to define a focus point of the excitation radiation

said fixing means (4) comprising adjustment means (8) for varying said focus point of the excitation radiation on a vertical axis to allow adjustment of the height of said focusing point within the chamber analysis.

Analysis chamber according to claim 1, characterized in that the second orifice is coincident with the first orifice.

Analysis chamber according to Claim 1 or 2, characterized in that the said fixing means (4) make it possible to move both the said light source and the said lens (22) for the convergence of the excitation radiation together. at the point of focus.

Analysis chamber according to claim 1 or 2, characterized in that said fixing means (4) enable said lens (22) to be moved, allowing the excitation radiation to converge at the focusing point, said light source being fixed by relative to the bottom wall of the analysis chamber.

Analysis chamber according to one of claims 1 to 4, characterized in that said fixing means (4) of said light source and said lens (22) includes a light-tight sleeve attached to the bottom wall of the chamber.

Analysis chamber according to one of claims 1 to 5, characterized in that said adjusting means (8) for varying the focus point of the excitation radiation comprise a millimeter screw.

A method of analyzing a sample present in an envelope comprising the steps of

(a) placing said sample on the floor of the chamber according to one of claims 1 to 6, vertically of said first orifice and facing said second orifice,

(c) emitting said excitation radiation and detecting Raman scattered light.

A method according to claim 7, characterized in that said sample is a liquid solution, and said envelope is selected from a syringe, an infusion bag, a bottle, a beaker or similar container, an ampoule, a portable infuser (comprising a first outer firm envelope containing a second flexible inner envelope containing said sample).

Process according to Claim 7, characterized in that the said sample is solid and the said envelope is chosen from a capsule, a capsule and a solid coating.

10. Method according to one of claims 8 to 9, characterized in that said excitation radiation is emitted for a period of between one second and one minute.